丝裂原活化蛋白激酶转导通路在葛根总黄酮诱导的急性早幼粒细胞白血病NB4细胞凋亡中的作用  

Effect of mitogen-activated protein kinase signaling pathway on apoptosis of acute promyelocytic leukemia NB4 cell line induced by puerariae radix flavones

作　　者：季鸥[1] 司叶俊 朱红青[2] 林琳[1] 姚浩 董雯 沈群[1] Ji Ou;Si Yejun;Zhu Hongqing;Lin Lin;Yao Hao;Dong Wen;Shen Qun(Department of Hematology,Affiliated Hospital of Nanjing University of Chinese Medicine,Nanjing 210029,China;the First Clinical College of Nanjing University of Chinese Medicine,Nanjing 210046,China)

机构地区：[1]南京中医药大学附属医院血液科,210029 [2]南京中医药大学第一临床医学院,210046

出　　处：《白血病．淋巴瘤》2020年第9期525-529,共5页Journal of Leukemia & Lymphoma

基　　金：国家自然科学基金(81703888、81274139)。

摘　　要：目的:探讨葛根总黄酮(PRF)诱导的急性早幼粒细胞白血病NB4细胞凋亡与丝裂原活化蛋白激酶(MAPK)转导通路相关信号分子的关系及意义。方法:将NB4细胞分为对照组[以0.025%二甲基亚砜(DMSO)代替PRF]和10、30、50μg/ml PRF组,作用24、48、72 h后用四甲基偶氮唑盐(MTT)法检测细胞增殖抑制率,作用48 h后用激光共聚焦显微镜观察细胞核形态学变化。将NB4细胞分为对照组(加入0.025%DMSO)和10、30、50μg/ml PRF联合或不联合10μmol/L c-Jun氨基末端激酶(JNK)抑制剂(SP600125)组,作用48 h,蛋白质印迹法检测MAPK通路相关蛋白JNK、肿瘤坏死因子α(TNF-α)、细胞外信号调节激酶(ERK)和p38 MAPK表达变化。结果:10、30、50μg/ml PRF作用24、48、72 h后NB4细胞增殖受到抑制,呈浓度-时间依赖性(均P<0.05),24、48、72 h半数抑制浓度(IC 50)分别为(40.03±2.23)μg/ml、(22.92±1.72)μg/ml、(17.99±1.48)μg/ml。激光共聚焦显微镜观察到NB4细胞经PRF处理后呈现明显凋亡特征。10μmol/L SP600125与不同浓度PRF共培养NB4细胞48 h后,NB4细胞JNK1表达受抑(P<0.05),JNK2/3、p38 MAPK表达有所下降,但差异均无统计学意义(均P>0.05);ERK1、ERK2在单药组表达逐渐上调,联合用药组则表达递减,TNF-α在50μg/ml PRF+SP600125组较50μg/ml PRF单药组表达下调,10、30μg/ml PRF+SP600125组则较10、30μg/ml PRF单药组表达上调。结论:10~50μg/ml PRF可能通过TNF-α激活MAPK信号途径,JNK、ERK1/2、p38 MAPK三者相互作用、相互影响,激活促凋亡相关蛋白,诱导NB4细胞凋亡。Objective To investigate the relationship between mitogen-activated protein kinase(MAPK)signaling pathway related signal molecules and the apoptosis of acute promyelocytic leukemia NB4 cells induced by puerariae radix flavones(PRF)and its significance.Methods The cells were divided into control group[0.025%dimethyl sulfoxide(DMSO)to replace PRF]and 10,30,50μg/ml PRF groups.The proliferation inhibition rate of NB4 cells exposed with PRF for 24,48 and 72 hours was determined by methyl thiazolyl tetrazolium(MTT)method,and the nuclear morphology was determined by confocal laser scanning microscope after 48 hours.NB4 cells were divided into control group(adding 0.025%DMSO)and 10,30 and 50μg/ml PRF with or without 10μmol/L c-Jun N-terminal kinase(JNK)inhibitor(SP600125)group,and the cells were treated for 48 hours and the changes in the expressions of MAPK pathway related proteins JNK,tumor necrosis factorα(TNF-α),extracellular signal-regulated kinase(ERK)and p38 MAPK were tested by Western blot.Results 10,30 and 50μg/ml PRF inhibited the proliferation of NB4 cells in 24,48 and 72 hours,which was in time-and dose-dependent manners(all P<0.05).The half-maximal inhibitory concentration(IC50)at 24,48 and 72 hours were(40.03±2.23)μg/ml,(22.92±1.72)μg/ml and(17.99±1.48)μg/ml,respectively.The confocal laser scanning microscope showed that NB4 cells displayed distinct apoptotic characteristics after PRF treatment.After co-cultivating NB4 cells with 10μmol/L SP600125 and different concentrations of PRF for 48 hours,the expression of JNK1 in NB4 cells was suppressed(P<0.05),and the expressions of JNK2/3 and p38 MAPK decreased,but the differences were not statistically significant(both P>0.05).The expressions of ERK1 and ERK2 gradually increased in the single-drug group,while the expression in the combined drug group decreased.The expression of TNF-αin the 50μg/ml PRF+SP600125 group was down-regulated compared with the 50μg/ml PRF single-drug group,while the expressions in the 10 and 30μg/ml PRF+SP600125 groups

关 键 词：白血病 早幼粒细胞 急性 葛根总黄酮 细胞凋亡 丝裂原活化蛋白激酶 

分 类 号：R733.71[医药卫生—肿瘤]